Crystal Structure of Haloalkane Dehalogenase LinB from Sphingomonas paucimobilis UT26 at 0.95 angstrom resolution.

Department of Biochemistry and Molecular Biology, Monash University* Australian National University** Masaryk University*** Palacky University**** Tohoku University*****
â—‹Matthew CJ Wilce* Aaron J Oakley** Martin Klvana*** Otyepka Michaal**** Yuji Nagata***** Jiri Damborsky***

Haloalkane dehalogenases utilize water to transform haloalkanes into organic halides and alcohols. The industrial production of halocarbons and the persistence of the compounds in the environment has led to interest in the enzymes for bioremediation purposes.

We have determined the crystal structure of LinB, a 33-kDa haloalkane dehalogenase from Sphingomonas paucimobilis UT26, at 0.95 Angstrom resolution. LinB belong to the alpha/beta hydrolase family of enzymes and consists of two domains.The data have allowed us to directly observe the anisotropic motions of the catalytic residues. In particular, the side-chain of the catalytic nucleophile, Asp108, displays a high degree of disorder. It has been modeled in two conformations, one similar to that observed previously (conformation A) and one strained (conformation B) that approached the catalytic base (His272). The strain in conformation B was mainly in the C(alpha)-C(beta)-C(gamma) angle (126 degrees ) that deviated by 13.4 degrees from the "ideal" bond angle of 112.6 degrees. On the basis of these observations, we propose a role for the charge state of the catalytic histidine in determining the geometry of the catalytic residues and that double-protonation of the catalytic base (His272) reduces the distance between the side-chain of this residue and that of the Asp108. Further, we were able to observe many alternative conformations of numerous side-chains and overlapping water networks.